It is well established that the multitude of microbes residing in the human intestine play a key role for health. Recently, it has become apparent that ingested chemicals affect the composition of the human gut microbiota. Additionally, the gut microbes affect the uptake and metabolism of chemicals in multiple ways. Here, we outline the current knowledge about the complex interplay between gut microbes, ingested xenobiotics and toxicological effects. We propose that the intestinal microbiota plays a key role in chemical toxicity, which is typically overlooked in existing approaches for risk assessment. This means that factors such as animal provider, batch/litter differences, and co-caging may significantly influence the outcome of toxicity evaluations based on rodent experiments.The effect of ingested chemicals may be significantly influenced by the gut microbiota of individual experimental animals. This should be considered in toxicological risk assessment.